PB1810: SELINEXOR COMBINED WITH CX-4945 PROMOTES CELL DEATH BY TARGETING PIK3CD/AKT1/FOXO3 SIGNALING IN ACUTE MYELOID LEUKEMIA
Authors: Huideng Long, Chunhua Song, Zheng Ge
Journal: HemaSphere (2023)

Topic: 3. Acute myeloid leukemia - Biology & Translational Research Background: Acute myeloid leukemia (AML) is a heterogeneous hematologic malignancy with high mortality and poor prognosis, which will always require highly effective treatment options. Selinexor (KPT-330) is a bioactive selective XPO1 inhibitor, and clinical trials have shown that the combination of KPT-330 is more beneficial to AML patients than standard induction therapy. CX-4945, an effective CK2 inhibitor, is reported to induce apoptosis and demonstrate therapeutic efficacy in high-risk leukemia. The antitumor effect of KPT-330 combined with CX-4945 has not been determined in AML. Aims: To explore the anti-leukemia effect of KPT-330 combined with CX-4945 in AML cells and its potential mechanism to provide evidence for clinical trials. Methods: Two AML patients’ BM samples were obtained from ZhongDa Hospital Southeast University, in which the cells were treated with indicated drugs in vitro. This study was approved by the Independent Ethics Committee for Clinical Research of Zhongda Hospital Southeast University. Cell Counting Kit-8 was used for cell viability assay, Annexin-V/PI staining following flow cytometry analysis for apoptosis, and qPCR for gene expression in AML cell lines and patient samples. CalcuSyn analysis was performed for a synergistic effect. RNA-seq was performed in U937 cells treated with 4mM CX-4945, the significantly differentially expressed genes (DEGs) were identified and gene set enrichment analysis (GSEA) was performed. Results: Cell proliferation arrest was significantly observed in U937 and MV4-11 cells treated with the combination of KPT-330 with CX-4945 compared to single drug control, and CalcuSyn analysis showed a synergistic effect of the combination on cell growth arrest (Fig. 1A). Then, the same trend was also noticed in 2 primary AML patients’ cells, and CalcuSyn analysis showed a synergistic effect of the combination on cell growth arrest (Fig.1B). Besides, the Total apoptosis rate increased significantly in the KPT-330+CX-4945 group in U937, MV-4-11, and primary AML patients’ cells compared to single drug controls (Fig.1C-1E). To understand the mechanisms underlying the synergy, RNA-seq analysis was performed in U937 cells treated by 4mM CX-4945 and 6979 DEGs were identified. PIK3CD and AKT1 are in the top DEGs upon CX-4945 treatment, GSEA and KEGG analysis indicated PI3K-AKT are enriched in DEGs (Fig. 1F-1H). XPO1 is the key component of nuclear-exporting machinery; FOXO3, the cargo of XPO1 is a translation initiation factor and plays a critical role in translational machinery. We observed a combination of KPT-330 with CX-4945 significantly down-regulated the mRNA level of oncogene PIK3CD or AKT1 and up-regulated the mRNA level of tumor suppressor gene FOXO3 in U937 and MV4-11 to single drug controls (Fig.1I). Moreover, the expression of AKT1 was increased and FOXO3 was decreased in AML patient cohort compared with healthy controls (Fig.1J); AKT1 was negatively correlated with FOXO3 in the AML patient cohort (Fig.1K), and the high expression of PIK3CD or AKT1 or low expression FOXO3 are associated with poor outcome (Fig.1L). These data indicated that PIK3CD/AKT1/FOXO3 signaling plays critical roles in the combination-mediated anti-leukemia, and the mechanistic model is summarized in Fig.1M. Summary/Conclusion: Our data demonstrated for the first time the synergistic effect of a novel combination of KPT-330 and CX-4945 on cell growth arrest and apoptosis in AML cells, and identify the underlying mechanism by targeting the PIK3CD/AKT1/FOXO3 signaling pathway, which highlighted the feasibility of clinical trials for combination therapy of AML patients.Keywords: AML